Background
Despite its effectiveness in suppressing active HIV replication, antiretroviral therapy (ART) does not eliminate the long-lived pool of HIV reservoir cells. Lymphoid tissues are a major reservoir even in the context of ART, but most studies on latent reservoirs focus on peripheral blood. The complex interplay between immune cells and the mechanisms that maintain viral persistence in lymphoid tissue remain incompletely understood. Advances in tissue imaging offer a unique opportunity to characterize the reservoir. Here, we used a spatial proteomic method to map immune microenvironments in HIV infected tissue.
Methods
Lymph node (LN) tissue from 6 people with HIV (PWH; n=3 undetectable, n=3 viremic) were obtained from the Pathology Anatomy Unit from Hospital Universitario La Princesa. LNs from people without HIV (PWOH) were included as negative controls (n=5). We applied MIBI-TOF (Multiplexed Ion Beam Imaging by Time of Flight), to simultaneously image 42 proteins in these tissues. We profiled over 1.8 million cells and characterized the spatial relationships between 20 immune cell types in their native tissue context. The inclusion of the viral protein p24 allowed us to identify p24+ and p24- microenvironments and their functional signatures.
Results
As expected, LNs from PWH had fewer CD4+ T cells compared to PWOH (p= 0.0006), while there were higher numbers of CD8+ T cells in both the whole tissue (p=0.024) and specifically within follicles (p=0.021). These CD8+ T cells expressed higher levels of Granzyme B and were enriched in p24+ LN (p=0.011). Further characterization of p24+ regions revealed higher levels of markers associated with activation and exhaustion of memory and effector CD8+ T cells (CD45RO, ICOS, TIGIT, Galectin9, TIM3), as well as inflammasome markers Caspase1 (p=0.03) and NLRP3 (p=0.3, FC=1.90). Using a new spatial niche detection method, we found that LN from PWH were enriched for CD8+ T cells near antigen-presenting cells (CD11c+, HLADR+), fibroblasts (SMA+), and endothelial cells (CD31+).
Conclusions
Using spatial proteomics to map the immune landscape in LNs, we identified novel markers characterizing CD8+ T cell subsets enriched in PWH with different levels of viremia, implying that active viral replication confers long-term changes on the immune landscape in LNs. Collectively, these data provide new insights into complex cell networks associated with viral replication at an important tissue reservoir site, which could be relevant for future HIV cure strategies.